The prototypic Fibroblast Growth Factors (FGFs), acidic FGF (also called FGF1) and basic FGF (also called FGF2) were first isolated in the 1970s (FGF2 by Gospodarowicz et al., J. Biol. Chem. 250:2515, 1975). There are currently 22 known FGF family members, which can be grouped into 7 subfamilies based upon their similarity in activities and sequences (Ornitz et al., Genome Biol. 2: 3005.1, 2001). However, the FGF family members bind to only four tyrosine kinase receptors (FGFR14) and their isoforms, which are expressed in a tissue-specific manner. The FGF 1 subgroup consists of FGF 1 and FGF2, which bind all four FGFRs, with FGF2 binding especially strongly to FGFR1c (Ornitz et al., J. Biol. Chem. 271:15292, 1996).
Human FGF2 is an 18 kDa non-glycosylated polypeptide consisting of 146 amino acids in the mature form derived from a 155 aa precursor (Ornitz et al., Genome Biol. 2:3005.1, 2001; Okada-Ban et al., Int. J. Biochem. Cell. Biol. 32:263, 2000). This 18 kDa form of FGF2 does not encode a signal sequence, but can be secreted by an unconventional energy-dependent pathway independent of the ER-Golgi complex (Mignatti et al., J. Cell Physiol. 151:81, 1992; Florkiewicz et al., J. Cell Physiol. 162:388, 1995). The single copy of the FGF2 gene also encodes four High Molecular Weight (HMW) forms of the protein, in addition to the 18 kDa form, by utilizing four alternate CUG initiation sites that provide N-terminal extensions of various sizes, resulting in proteins of 22 kDa (196 aa), 22.5 kDa (201 aa), 24 kDa (210 aa) and 34 kDa (288 aa) (Florkiewicz et al., Proc. Natl. Acad. Sci. USA 86:3978, 1989; Prats et al., Proc. Natl. Acad. Sci. USA 86:1836, 1989). The HMW forms are not secreted but are transported to the cell nucleus where they can regulate cell growth or behavior in an intracrine fashion (Delrieu, FEBS Lett. 468:6, 2000).
In addition to binding FGFR1-4 with high affinity, FGF2 binds to heparin sulfate proteoglycans (HSPG) with lower affinity. Although FGF2 is secreted as a monomer, cell surface HSPG dimerizes FGF2 in a non-covalent side-to-side configuration that is subsequently capable of dimerizing and activating FGF receptors (Mohammadi et al., Cytokine Growth Factor Rev 16:107, 2005). The binding of FGF and HSPG to the extracellular domain of FGFR induces receptor dimerization, activation and autophosphorylation.
The FGFs, and in particular FGF2, have a broad spectrum of activities on various cell types (Ornitz et al., Genome Biol. 2:3005.1, 2001). FGF2 stimulates proliferation of (i.e., is mitogenic for) certain cells including fibroblasts and endothelial cells and is a survival factor (anti-apoptotic) for certain cells such as neural cells (Okada-Ban, op. cit.). It also stimulates differentiation (morphogenesis) and migration (motility) of endothelial cells (Dow et al., Urology 55:800, 2000). FGF2 is involved in development, especially of the nervous system. Importantly, FGF2 is a powerful angiogenic factor (Presta et al., Cytokine and Growth Factor Rev. 16:159, 2005).
FGF2 and other FGFs are believed to play a role in cancer, both by stimulating angiogenesis and tumor cells directly (Presta et al., op cit.) During tumor progression, cancer cells may respond to the extracellular FGF2 secreted from the stromal cells (paracrine), and then the tumor cells themselves may secrete FGF2 and respond to it in an autocrine manner. FGF2 or its receptor FGFR1 has been shown to be expressed or overexpressed in most gliomas (Takahashi et al., Proc. Natl. Acad. Sci. USA 87:5710, 1990; Morrison et al. Cancer Res. 54:2794, 1994). FGF2 is involved in progression of prostate tumors (Dow et al., Urology 55:800, 2000) and is a key mediator of the proliferation of malignant melanomas (Wang et al., Nature Med. 3:887, 1997). Over-expression and/or involvement of FGF2 or FGFR1 in tumor progression has also been reported for salivary gland tumors (Myoken et al., J. Path. 178:429, 1996), esophageal cancer (Barclay et al., Clin. Cancer Res. 11:7683, 2005), and thyroid carcinomas (Boelaert et al., J. Clin. Endocrin. Metabol. 88:2341, 2003).
Polyclonal antibodies (antiserum) to FGF2 have been reported to inhibit tumor growth of a transplantable chondrosarcoma in mice (Baird et al., J. Cell Biochem. 30:79, 1986), neutralize various activities of FGF2 in vitro (Kurokawa et al., J. Biol. Chem. 264:7686, 1989), and to inhibit growth of U87 glioma cell intracranial xenografts when applied locally (Stan et al., J. Neurosurg. 82:1044, 1995). Among monoclonal antibodies, the DG2 mAb has been reported to neutralize activities of FGF2 in vitro and in vivo (Reilly et al., Biochem. Biophys. Res. Com. 164:736, 1989; WO 91/06668), modestly inhibit growth of rat C6 glioma xenografts in nude mice (Gross et al., J. Nat. Cancer Inst. 85:121, 1993), and modestly inhibit growth of rat chondrosarcomas when delivered intralesionally but not i.p. or i.v. (Coppola et al., Anticancer Res. 17:2033, 1997). Similarly, anti-FGF2 mAb DE6 has been reported to inhibit growth of glioma cells in vitro (Morrison et al., J. Neuroscience Res. 34:502, 1993). On the other hand, the anti-FGF2 mAbs bFM-1 and bFM-2 was reported to inhibit growth of endothelial cells in vitro but not to block tumor angiogenesis in vivo (Matsuzaki et al., Proc. Natl. Acad. Sci. USA 86:9911, 1989). The neutralizing anti-FGF2 mAb 1E6 has been reported to inhibit growth of RPMI4788 colon tumor xenografts (Aonuma et al., 19:4039, 1999). The anti-FGF2 mAb 254F1 has been reported to inhibit proliferation of human umbilical vein endothelial cells (HUVEC), while mAb FB-8 has been reported to inhibit proliferation of non-small cell lung cancer cell lines in vitro (Kuhn et al., Lung Cancer 44:167, 2004). The anti-FGF2 mAb 3H3 was reported to suppress growth of U87MG and T98G glioma and HeLa cell xenografts (Takahashi et al., FEBS Let. 288:65, 1991) and growth of the K1000 FGF2-transfected 3T3 cell line in mice (Hori et al., Cancer Res. 51:6180, 1991).
The GAL-F2 anti-FGF2 mAb described herein was discussed at the AACR 2009 Annual Meeting in Poster #1236, which is incorporated herein in its entirety for all purposes.